The major aim of the proposed study is to investigate fundamental mechanisms of electromechanical coupling in smooth muscle using the canine sphincter of Oddi (SO) as a model. The sphincter of Oddi plays a pivotal role in control of the outflow of bile and pancreatic juices into the duodenum. This is achieved by rhythmic changes in smooth muscle tone through changes in membrane potential of the smooth muscle. Smooth muscle membrane potential is controlled by (a) myogenic activity inherent to the smooth muscle, and (b) by inputs from excitatory cholinergic nerves, inhibitory non-adrenergic non-cholinergic nerves (NANC), and by the enteric hormone, cholecystokinin. Firstly, I will investigate the ionic basis of electrical myogenic rhythmicity in the smooth muscle by characterizing the major ionic currents expressed in smooth muscle cells isolated from the sphincter of Oddi. I will explore intra-SO regional differences in the expression of "pacemaker" current, K-channel currents and Ca-channel current. Secondly I will investigate the ionic mechanisms by which acetylcholine (ACh), a major excitatory neurotransmitter in the SO, excites the smooth muscle and whether the channels activated by ACh conduct Ca2+. I will then investigate the ionic mechanisms and intracellular second messenger pathways by which the inhibitory actions of vasoactive intestinal polypeptide (VIP), a putative NANC inhibitory neurotransmitter in the SO, are transduced. I will test that the inhibition produced by both VIP and the NANC inhibitory neurotransmitter occurs by the opening of ATP-sensitive K+ channels in the smooth muscle. I will test the direct action of cholecystokinin (CCK) on the smooth muscle of the SO by characterizing the ionic current activated by this enteric hormone. Finally, because electrical rhythmicity determines the pattern of contractile activity, I will investigate the direct relationship between membrane potential, voltage-gated Ca2+ channel current and the cytoplasmic [Ca2+]. The experiments to be conducted in this study will employ advanced electrophysiological techniques to voltage-clamp single cells and record ionic currents from many channels and from individual ionic channels, and simultaneous voltage-clamp and ratiometric photometry of cytoplasmic [Ca2+] using Ca2+-sensitive dyes. In addition conventional intracellular microelectrode recordings will be employed on multicellular smooth muscle strips from the SO. Motor diseases of the SO affect contractile rhythmicity and the resistance to flow of the SO. The resultant impediment to outflow of bile and pancreatic enzymes has serious effects on health. By studying the fundamental mechanisms by which motility in the SO is generated and modulated, we will gain a clearer understanding of the etiology of these disorders and be able to develop specific therapeutic agents.